This study has been concerned with the development and establishment of endothelial cell cultures derived from three microvascular fractions of human brain. The purity of the endothelial cultures was found to be greater than 95% since most of the cells expressed human Factor VIII-related antigen and only an occasional cell stained for glial fibrillary acidic protein. Both light and electron microscopy showed that the separately cultured endothelium derived either from the capillaries (EC) or small [(<100mu-m)ES] and large [>100mu-m) EL] arterioles and venules exhibited the same appearance and growth pattern. The maximal proliferative response of endothelium was seen on the first day after exposure to fresh medium. A direct relationship was found between various components of growth medium and individual proliferative response of endothelium dependent on its origin indicating different growth requirements. The characterization of the cultured endothelium is still incomplete and in progress. Nevertheless the successfully developed technique for cultivating pure cerebromicrovascular endothelium derived from human brain (dependent on the availability of brain tissue) provides a model in vitro for investigation of endothelial properties related to the normal or altered function of blood-brain barrier and cerebral blood flow in man. The loss of endothelial cultures due to toxic human serum prevented the completion of this project.